The aim of this project is to identify, characterize, and clone those genes that drive the development of neoplasia and whose malignant potential results from changes caused by chemical carcinogens. (1) DNA from three BALB/3T3 cell lines transformed by benzo[a]pyrene (BP) was analyzed by DNA transfer and focus formation in the NIH/3T3 system. All three tested lines showed transforming activity that differed from each other and from the ras oncogenes by restriction endonuclease sensntivity and MspI mapping. These possibly new transforming genes are now being cloned by the sib selection protocol from Charon 4A phage genomic libraries. Several cycles of selections have been carried out with oncogene activity and the positive pools show no presence of the ras oncogenes. (2) The mechanism by which chemical carcinogens may activate proto-oncogenes was explored by analyzing the distribution of carcinogen adducts on different parts of the genome. BP adducts, formed in vivo in hamster liver cells were preferentially located in DNAse I hypersensitive regions of the genome and were rapidly removed by repair processes, while persisting adducts remained in other parts of the genome. By this approach, the Ha-ras proto-oncogene was found to be present in a transcriptionally active form in target hamster liver cells. (3) Human homologs of pro genes were isolated from the human nasapharyngeal carcinoma cell line CNE2, and were characterized.
